Skip to main content

Advertisement

Log in

Rapid Shifts in the Structure and Composition of a Protistan Assemblage During Bottle Incubations Affect Estimates of Total Protistan Species Richness

  • Microbiology of Aquatic Systems
  • Published:
Microbial Ecology Aims and scope Submit manuscript

Abstract

Changes in the structure and composition of a protistan community were characterized through the analysis of small-subunit ribosomal RNA gene (18S) sequences for a 3-day bottle incubation using a single sample collected in the western North Atlantic. Cloning and sequencing was used to investigate changes in perceived species richness and diversity as a consequence of environmental perturbation. The treatments included a control (unamended seawater), inorganic nutrient enrichment, and enrichment with a complex organic mixture. Five clone libraries were constructed and analyzed at the time of collection (t-0 h) and after 24 (t-24 h) and 72 (t-72 h) h for the control, and at t-72 h for the inorganic and organic enrichments, resulting in an analysis of 1,626 partial 18S rDNA sequences that clustered into 238 operational taxonomic units (OTUs). Analysis of the clone libraries revealed that protistan assemblages were highly dynamic and changed substantially at both the OTU level and higher taxonomic classifications during time frames consistent with many oceanographic methods used for measuring biological rates. Changes were most dramatic in enrichments, which yielded community compositions that were strongly dominated by one or a few taxa. Changes in community structure during incubation dramatically influenced estimates of species richness, which were substantially lower with longer incubation and especially with amendment, even though all incubated samples originated from the same aliquot of seawater. Containment and enrichment of the seawater sample led to the detection of otherwise undetected protistan taxa, suggesting that characterization of protistan diversity in a sample only at the time of collection could lead to an underrepresentation of unique taxa. Additionally, the rapid increase in the relative abundance of some members of the “rare biosphere” in our results implies an ecological importance of at least some of the taxa comprising the “rare biosphere.”

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

References

  1. Agis M, Granda A, Dolan JR (2007) A cautionary note: examples of possible microbial community dynamics in dilution grazing experiments. J Exp Mar Biol Ecol 341:176–183

    Article  Google Scholar 

  2. Aguilera A, Souza-Egipsy V, Gonzalez-Toril E, Rendueles O, Amils R (2010) Eukaryotic microbial diversity of phototrophic microbial mats in two Icelandic geothermal hot springs. Int Microbiol 13:21–32

    PubMed  CAS  Google Scholar 

  3. Altschul SF, Madden TL, Schaffer AA, Zhang JH, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  PubMed  CAS  Google Scholar 

  4. Ashelford KE, Chuzhanova NA, Fry JC, Jones AJ, Weightman AJ (2005) At least 1 in 20 16 S rRNA sequence records currently held in public repositories is estimated to contain substantial anomalies. Appl Environ Microbiol 71:7724–7736

    Article  PubMed  CAS  Google Scholar 

  5. Barbeau K, Moffett JW, Caron DA, Croot PL, Erdner DL (1996) Role of protozoan grazing in relieving iron limitation of phytoplankton. Nature 380:61–64

    Article  CAS  Google Scholar 

  6. Bray JR, Curtis JT (1957) An ordination of the upland forest communities of Southern Wisconsin. Ecol Monogr 27:326–349

    Article  Google Scholar 

  7. Bunge J, Barger K (2008) Parametric models for estimating the number of classes. Biom J 50:971–982

    Article  PubMed  Google Scholar 

  8. Caron DA (2000) Protistan herbivory and bacterivory. Method Microbiol 30(30):289–315

    Google Scholar 

  9. Caron DA (2009) New accomplishments and approaches for assessing protistan diversity and ecology in natural ecosystems. BioSci 59:287–299

    Article  Google Scholar 

  10. Caron DA, Countway PD (2009) Hypotheses on the role of the protistan rare biosphere in a changing world. Aquat Microb Ecol 57:227–238

    Article  Google Scholar 

  11. Caron DA, Countway PD, Brown MV (2004) The growing contributions of molecular biology and immunology to protistan ecology: molecular signatures as ecological tools. J Eukaryot Microbiol 51:38–48

    Article  PubMed  CAS  Google Scholar 

  12. Caron DA, Countway PD, Savai P, Gast RJ, Schnetzer A, Moorthi SD, Dennett MR, Jones AC (2009) Defining DNA-based operational taxonomic units for microbial-eukaryote ecology. Appl Environ Microbiol 75:5797–5808

    Article  PubMed  CAS  Google Scholar 

  13. Caron DA, Dennett MR, Lonsdale DJ, Moran DM, Shalapyonok L (2000) Microzooplankton herbivory in the Ross Sea, Antarctica. Deep Sea Res (II Top Stud Oceanogr) 47:3249–3272

    Article  Google Scholar 

  14. Chao A, Shen T-J (2010) Program SPADE (Species Prediction And Diversity Estimation). Program and User’s Guide. Available at: http://chao.stat.nthu.edu.tw

  15. Cole JR, Chai B, Marsh TL, Farris RJ, Wang Q, Kulam SA, Chandra S, McGarrell DM, Schmidt TM, Garrity GM, Tiedje JM (2003) The Ribosomal Database Project (RDP-II): previewing a new autoaligner that allows regular updates and the new prokaryotic taxonomy. Nucleic Acids Res 31:442–443

    Article  PubMed  CAS  Google Scholar 

  16. Collado-Mercado E, Radway JC, Collier JL (2010) Novel uncultivated labyrinthulomycetes revealed by 18 S rDNA sequences from seawater and sediment samples. Aquat Microb Ecol 58:215–228

    Article  Google Scholar 

  17. Countway PD, Gast RJ, Savai P, Caron DA (2005) Protistan diversity estimates based on 18 S rDNA from seawater incubations in the western North Atlantic. J Eukaryot Microbiol 52:95–106

    Article  PubMed  CAS  Google Scholar 

  18. Dawson SC, Pace NR (2002) Novel kingdom-level eukaryotic diversity in anoxic environments. Proc Natl Acad Sci USA 99:8324–8329

    Article  PubMed  CAS  Google Scholar 

  19. Diez B, Pedros-Alio C, Massana R (2001) Study of genetic diversity of eukaryotic picoplankton in different oceanic regions by small-subunit rRNA gene cloning and sequencing. Appl Environ Microbiol 67:2932–2941

    Article  PubMed  CAS  Google Scholar 

  20. Dolan JR, McKeon K (2004) The reliability of grazing rate estimates from dilution experiments: have we over-estimated rates of organic carbon consumption by microzooplankton? Ocean Sci 1:1–7

    Article  Google Scholar 

  21. Doney SC (2010) The growing human footprint on coastal and open-ocean biogeochemistry. Science 328:1512–1516

    Article  PubMed  CAS  Google Scholar 

  22. Dopheide A, Lear G, Stott R, Lewis G (2009) Relative diversity and community structure of ciliates in stream biofilms according to molecular and microscopy methods. Appl Environ Microbiol 75:5261–5272

    Article  PubMed  CAS  Google Scholar 

  23. Edgcomb VP, Kysela DT, Teske A, Gomez AD, Sogin ML (2002) Benthic eukaryotic diversity in the Guaymas Basin hydrothermal vent environment. Proc Natl Acad Sci USA 99:7658–7662

    Article  PubMed  CAS  Google Scholar 

  24. Eppley RW (1968) An incubation method for estimating carbon content of phytoplankton in natural samples. Limnol Oceanogr 13:574

    Article  Google Scholar 

  25. Epstein S, Lopez-Garcia P (2008) “Missing” protists: a molecular prospective. Biodivers Conserv 17:261–276

    Article  Google Scholar 

  26. Hughes JB, Hellmann JJ (2005) The application of rarefaction techniques to molecular inventories of microbial diversity. Method Enzymol 397:292–308

    Article  CAS  Google Scholar 

  27. Hutchins DA, Bruland KW (1998) Iron-limited diatom growth and Si: N uptake ratios in a coastal upwelling regime. Nature 393:561–564

    Article  CAS  Google Scholar 

  28. Hutchins DA, Mulholland MR, Fu FX (2009) Nutrient cycles and marine microbes in a CO2-enriched ocean. Oceanography 22:128–145

    Google Scholar 

  29. Kolodziej K, Stoeck T (2007) Cellular identification of a novel uncultured marine stramenopile (MAST-12 clade) small-subunit rRNA gene sequence from a Norwegian estuary by use of fluorescence in situ hybridization-scanning electron microscopy. Appl Environ Microbiol 73:2718–2726

    Article  PubMed  CAS  Google Scholar 

  30. Kruskal W, Kruskal W (1967) Statistics, Moliere, and Henry Adams. Am Sci 55:416

    Google Scholar 

  31. Landry MR, Kirshtein J, Constantinou J (1995) A refined dilution technique for measuring the community grazing impact of microzooplankton, with experimental tests in the Central Equatorial Pacific. Mar Ecol Prog Ser 120:53–63

    Article  Google Scholar 

  32. Lomas MW, Bates NR (2004) Potential controls on interannual partitioning of organic carbon during the winter/spring phytoplankton bloom at the Bermuda Atlantic Time-series Study (BATS) site. Deep Sea Res (I-Oceanogr Res Papers) 51:1619–1636

    CAS  Google Scholar 

  33. Lopez-Garcia P, Rodriguez-Valera F, Pedros-Alio C, Moreira D (2001) Unexpected diversity of small eukaryotes in deep-sea Antarctic plankton. Nature 409:603–607

    Article  PubMed  CAS  Google Scholar 

  34. Lovejoy C, Massana R, Pedros-Alio C (2006) Diversity and distribution of marine microbial eukaryotes in the Arctic Ocean and adjacent seas. Appl Environ Microbiol 72:3085–3095

    Article  PubMed  CAS  Google Scholar 

  35. Massana R, Castresana J, Balague V, Guillou L, Romari K, Groisillier A, Valentin K, Pedros-Alio C (2004) Phylogenetic and ecological analysis of novel marine stramenopiles. Appl Environ Microbiol 70:3528–3534

    Article  PubMed  CAS  Google Scholar 

  36. Massana R, Guillou L, Diez B, Pedros-Alio C (2002) Unveiling the organisms behind novel eukaryotic ribosomal DNA sequences from the ocean. Appl Environ Microbiol 68:4554–4558

    Article  PubMed  CAS  Google Scholar 

  37. Massana R, Pedros-Alio C, Casamayor EO, Gasol JM (2001) Changes in marine bacterioplankton phylogenetic composition during incubations designed to measure biogeochemically significant parameters. Limnol Oceanogr 46:1181–1188

    Article  Google Scholar 

  38. Massana R, Terrado R, Forn I, Lovejoy C, Pedros-Alio C (2006) Distribution and abundance of uncultured heterotrophic flagellates in the world oceans. Environ Microbiol 8:1515–1522

    Article  PubMed  CAS  Google Scholar 

  39. McGradySteed J, Harris PM, Morin PJ (1997) Biodiversity regulates ecosystem predictability. Nature 390:162–165

    Article  CAS  Google Scholar 

  40. Moon-van der Staay SY, De Wachter R, Vaulot D (2001) Oceanic 18 S rDNA sequences from picoplankton reveal unsuspected eukaryotic diversity. Nature 409:607–610

    Article  PubMed  CAS  Google Scholar 

  41. Moreira D, Lopez-Garcia P (2002) The molecular ecology of microbial eukaryotes unveils a hidden world. Trends Microbiol 10:31–38

    Article  PubMed  CAS  Google Scholar 

  42. Moreira D, Lopez-Garcia P (2003) Are hydrothermal vents oases for parasitic protists? Trends Parasitol 19:556–558

    Article  PubMed  CAS  Google Scholar 

  43. Naeem S, Li SB (1997) Biodiversity enhances ecosystem reliability. Nature 390:507–509

    Article  CAS  Google Scholar 

  44. Not F, del Campo J, Balague V, de Vargas C, Massana R (2009) New insights into the diversity of marine picoeukaryotes. PLoS ONE 4:e7143

    Article  PubMed  Google Scholar 

  45. Not F, Valentin K, Romari K, Lovejoy C, Massana R, Tobe K, Vaulot D, Medlin LK (2007) Picobiliphytes: a marine picoplanktonic algal group with unknown affinities to other eukaryotes. Science 315:253–255

    Article  PubMed  CAS  Google Scholar 

  46. Ohman MD, Snyder RA (1991) Growth-kinetics of the omnivorous oligotrich ciliate Strombidium sp. Limnol Oceanogr 36:922–935

    Article  Google Scholar 

  47. Pace NR, Stahl DA, Lane DJ, Olsen GJ (1986) The analysis of natural microbial-populations by ribosomal-RNA sequences. Adv Microb Ecol 9:1–55

    CAS  Google Scholar 

  48. Pedros-Alio C (2007) Dipping into the rare biosphere. Science 315:192–193

    Article  PubMed  CAS  Google Scholar 

  49. Potvin M, Lovejoy C (2009) PCR-based diversity estimates of artificial and environmental 18 S rRNA gene libraries. J Eukaryot Microbiol 56:174–181

    Article  PubMed  CAS  Google Scholar 

  50. Rodriguez-Martinez R, Labrenz M, del Campo J, Forn I, Jurgens K, Massana R (2009) Distribution of the uncultured protist MAST-4 in the Indian Ocean, Drake Passage and Mediterranean Sea assessed by real-time quantitative PCR. Environ Microbiol 11:397–408

    Article  PubMed  CAS  Google Scholar 

  51. Rose JM, Caron DA (2007) Does low temperature constrain the growth rates of heterotrophic protists? Evidence and implications for algal blooms in cold waters. Limnol Oceanogr 52:886–895

    Article  Google Scholar 

  52. Sherr BF, Sherr EB, Caron DA, Vaulot D, Worden AZ (2007) Oceanic protists. Oceanography 20:130–134

    Google Scholar 

  53. Sherr EB, Sherr BF (1994) Bacterivory and herbivory—key roles of phagotrophic protists in pelagic food webs. Microb Ecol 28:223–235

    Article  Google Scholar 

  54. Sherr EB, Sherr BF (2002) Significance of predation by protists in aquatic microbial food webs. Antonie Van Leeuwenhoek Intl J Gen Mol Microbiol 81:293–308

    Article  CAS  Google Scholar 

  55. Shi XL, Marie D, Jardillier L, Scanlan DJ, Vaulot D (2009) Groups without cultured representatives dominate eukaryotic picophytoplankton in the oligotrophic South East Pacific Ocean. PLoS ONE 4:e7657

    Article  PubMed  Google Scholar 

  56. Sogin ML, Morrison HG, Huber JA, Welch DM, Huse SM, Neal PR, Arrieta JM, Herndl GJ (2006) Microbial diversity in the deep sea and the underexplored “rare biosphere”. Proc Natl Acad Sci USA 103:12115–12120

    Article  PubMed  CAS  Google Scholar 

  57. Stepanauskas R, Sieracki ME (2007) Matching phylogeny and metabolism in the uncultured marine bacteria, one cell at a time. Proc Natl Acad Sci USA 104:9052–9057

    Article  PubMed  CAS  Google Scholar 

  58. Stoeck T, Epstein S (2003) Novel eukaryotic lineages inferred from small-subunit rRNA analyses of oxygen-depleted marine environments. Appl Environ Microbiol 69:2657–2663

    Article  PubMed  CAS  Google Scholar 

  59. Stoecker DK, Johnson MD, de Vargas C, Not F (2009) Acquired phototrophy in aquatic protists. Aquat Microb Ecol 57:279–310

    Article  Google Scholar 

  60. Strong A, Chisholm S, Miller C, Cullen J (2009) Ocean fertilization: time to move on. Nature 461:347–348

    Article  PubMed  CAS  Google Scholar 

  61. Takishita K, Yubuki N, Kakizoe N, Inagaki Y, Maruyama T (2007) Diversity of microbial eukaryotes in sediment at a deep-sea methane cold seep: surveys of ribosomal DNA libraries from raw sediment samples and two enrichment cultures. Extremophiles 11:563–576

    Article  PubMed  CAS  Google Scholar 

  62. Tekle YI, Parfrey LW, Katz LA (2009) Molecular data are transforming hypotheses on the origin and diversification of eukaryotes. BioSci 59:471–481

    Article  Google Scholar 

  63. Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W—improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680

    Article  PubMed  CAS  Google Scholar 

  64. Vigil P, Countway PD, Rose J, Lonsdale DJ, Gobler CJ, Caron DA (2009) Rapid shifts in dominant taxa among microbial eukaryotes in estuarine ecosystems. Aquat Microb Ecol 54:83–100

    Article  Google Scholar 

  65. Watt WD (1971) Measuring primary production rates of individual phytoplankton species in natural mixed populations. Deep-Sea Res 18:329

    Google Scholar 

  66. Weekers PHH, Gast RJ, Fuerst PA, Byers TJ (1994) Sequence variations in small-subunit ribosomal-RNAs of Hartmannella vermiformis and their phylogenetic implications. Mol Biol Evol 11:684–690

    PubMed  CAS  Google Scholar 

  67. Worden AZ (2006) Picoeukaryote diversity in coastal waters of the Pacific Ocean. Aquat Microb Ecol 43:165–175

    Article  Google Scholar 

  68. Youssef NH, Couger MB, Elshahed MS (2010) Fine-scale bacterial beta diversity within a complex ecosystem (Zodletone Spring, OK, USA): the role of the rare biosphere. PLoS ONE 5:e12414

    Article  PubMed  Google Scholar 

  69. Youssef NH, Elshahed MS (2008) Species richness in soil bacterial communities: a proposed approach to overcome sample size bias. J Microbiol Meth 75:86–91

    Article  Google Scholar 

  70. Zobell CE (1943) The effect of solid surfaces upon bacterial activity. J Bacteriol 46:39–56

    PubMed  CAS  Google Scholar 

  71. Zuendorf A, Bunge J, Behnke A, Barger KJA, Stoeck T (2006) Diversity estimates of microeukaryotes below the chemocline of the anoxic Mariager Fjord, Denmark. FEMS Microbiol Ecol 58:476–491

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This study was supported by the National Science Foundation grants OCE-9818953, MCB-0084231, OCE-0550829, and MCB-0703159. The authors would like to thank the following people for assistance with this study: Mark Dennett, Dawn Moran, David Beaudoin, Rebecca Shaffner, and Julie Rose for their participation on the research cruise aboard R/V Endeavor and the captain and crew of the R/V Endeavor.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Diane Y. Kim.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kim, D.Y., Countway, P.D., Gast, R.J. et al. Rapid Shifts in the Structure and Composition of a Protistan Assemblage During Bottle Incubations Affect Estimates of Total Protistan Species Richness. Microb Ecol 62, 383–398 (2011). https://doi.org/10.1007/s00248-011-9816-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00248-011-9816-9

Keywords

Navigation